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Overexpression of klotho in adipose-derived stem cells protects against UVB-induced photoaging in co-cultured human fibroblasts.

ABSTRACT: Co-culture methods are widely used in tissue engineering to drive tissue formation with the direct or indirect interaction of multiple cell types. Klotho is a novel biomarker involved in aging. In this study, we evaluated the protective effects of klotho overexpressed adipose-derived stem cells (ADSCs) against ultraviolet radiation B (UVB)-induced photoaging in co-cultured human skin fibroblasts (HSF2 cell line). Furthermore, the involvement of P38 mitogen-activated protein kinase (MAPK) signaling was investigated. ADSCs were isolated from human subcutaneous adipose tissue and the 3rd generation of ADSCs was used after being identified. Klotho overexpression (OE) lentivirus vectors were constructed and identified in ADSCs. The HSF2 cells were seeded in the upper layer of the Transwell co-culture plate (0.4 µm pore polycarbonate membrane) and ADSCs were seeded in the lower layer. UVB irradiation of HSF2 cells was performed using UVB lamps in uncovered petri dishes at room temperature. The present results indicated that the proliferation of ADSCs was increased by klotho OE. Furthermore the proliferation and collagen content of HSF2 were decreased by UVB irradiation in a dose-dependent manner. By contrast, the protein level of matrix metalloproteinases (MMP) 1, 3 and p-P38 in HSF2 were upregulated. In the co-culture system, relative mRNA expression of MMP-1 and MMP-3 as well as protein level of MMP-1, MMP-3 and p-P38 in HSF2 were reduced by co-culture with klotho overexpressed ADSCs when exposed to UVB (20 mJ/cm2). By contrast, the collagen content of HSF2 was increased. Collectively, OE of klotho in ADSCs notably ameliorates UVB-induced photoaging in co-cultured HSF2, and these effects were potentially achieved by increasing the collagen content and decreasing the protein level of MMP-1, MMP-3 and p-P38.

SUBMITTER: Fan F

PROVIDER: S-EPMC6236291 | biostudies-literature | 2018 Dec

REPOSITORIES: biostudies-literature

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Overexpression of klotho in adipose-derived stem cells protects against UVB-induced photoaging in co-cultured human fibroblasts.

Fan Furong F Li Yufei Y Liu Yi Y Shao Li L Yu Juehua J Li Ziwei Z

Molecular medicine reports 20181025 6

Co-culture methods are widely used in tissue engineering to drive tissue formation with the direct or indirect interaction of multiple cell types. Klotho is a novel biomarker involved in aging. In this study, we evaluated the protective effects of klotho overexpressed adipose-derived stem cells (ADSCs) against ultraviolet radiation B (UVB)-induced photoaging in co-cultured human skin fibroblasts (HSF2 cell line). Furthermore, the involvement of P38 mitogen-activated protein kinase (MAPK) signali ...[more]

PMID: 30365106

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Project description:Skin photoaging due to ultraviolet B (UVB) exposure generates reactive oxygen species (ROS) that increase matrix metalloproteinase (MMP). Chlorin e6-photodynamic therapy (Ce6-PDT), in addition to being the first-line treatment for malignancies, has been shown to lessen skin photoaging, while curcumin is well known for reducing the deleterious effects of ROS. In the current study, PDT with three novel Ce6-curcumin derivatives, a combination of Ce6 and curcumin with various linkers, including propane-1,3-diamine for Ce6-propane-curcumin; hexane-1,6-diamine for Ce6-hexane-curcumin; and 3,3'-((oxybis(ethane-2,1-diyl))bis(oxy))bis(propan-1-amine) for Ce6-dipolyethylene glycol (diPEG)-curcumin, were studied for regulation of UVB-induced photoaging on human skin fibroblast (Hs68) and mouse embryonic fibroblast (BALB/c 3T3) cells. We assessed the antiphotoaging effects of Ce6-curcumin derivatives on cell viability, antioxidant activity, the mechanism of matrix metalloproteinase-1 and 2 (MMP-2) expression, and collagen synthesis in UVB-irradiated in vitro models. All three Ce6-curcumin derivatives were found to be non-phototoxic in the neutral red uptake phototoxicity test. We found that Ce6-hexane-curcumin-PDT and Ce6-propane-curcumin-associated PDT exhibited less cytotoxicity in Hs68 and BALB/c 3T3 fibroblast cell lines compared to Ce6-diPEG-curcumin-PDT. Ce6-diPEG-curcumin and Ce6-propane-curcumin-associated PDT showed superior antioxidant activity in Hs68 cell lines. Further, in UVB-irradiated in vitro models, the Ce6-diPEG-curcumin-PDT greatly attenuated the expression levels of MMP-1 and MMP-2 by blocking mitogen-activated protein kinases (MAPKs), activator protein 1 (AP-1), and tumor necrosis factor-α (NF-κB) signaling. Moreover, Ce6-diPEG-curcumin effectively inhibited inflammatory molecules, such as cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression, while accelerating collagen synthesis. These results demonstrate that Ce6-diPEG-curcumin may be a potential therapy for treating skin photoaging.

Project description:A number of natural phytochemicals have anti-photoaging properties that appear to be mediated through the inhibition of matrix metalloproteinase-1 (MMP-1) expression, but their direct target molecule(s) and mechanism(s) remain unclear. We investigated the effect of naringenin, a major flavonoid found in citrus, on UVB-induced MMP-1 expression and identified its direct target. The HaCaT human skin keratinocyte cell line and 3-dimensional (3-D) human skin equivalent cultures were treated or not treated with naringenin for 1 hr before exposure to UVB. The mechanism and target(s) of naringenin were analysed by kinase assay and multiplex molecular assays. Dorsal skins of hairless mice were exposed to UVB 3 times per week, with a dose of irradiation that was increased weekly by 1 minimal erythema dose (MED; 45 mJ/cm(2)) to 4 MED over 15 weeks. Wrinkle formation, water loss and water content were then assessed. Naringenin suppressed UVB-induced MMP-1 expression and AP-1 activity, and strongly suppressed UVB-induced phosphorylation of Fos-related antigen (FRA)-1 at Ser265. Importantly, UVB irradiation-induced FRA1 protein stability was reduced by treatment with naringenin, as well as with a mitogen-activated protein kinase (MEK) inhibitor. Naringenin significantly suppressed UVB-induced extracellular signal-regulated kinase 2 (ERK2) activity and subsequently attenuated UVB-induced phosphorylation of p90(RSK) by competitively binding with ATP. Constitutively active MEK (CA-MEK) increased FRA1 phosphorylation and expression and also induced MMP-1 expression, whereas dominant-negative ERK2 (DN-ERK2) had opposite effects. U0126, a MEK inhibitor, also decreased FRA1 phosphorylation and expression as well as MMP-1 expression. The photoaging data obtained from mice clearly demonstrated that naringenin significantly inhibited UVB-induced wrinkle formation, trans-epidermal water loss and MMP-13 expression. Naringenin exerts potent anti-photoaging effects by suppressing ERK2 activity and decreasing FRA1 stability, followed by down-regulation of AP-1 transactivation and MMP-1 expression.

Dataset Information

Overexpression of klotho in adipose-derived stem cells protects against UVB-induced photoaging in co-cultured human fibroblasts.

Publications

Overexpression of klotho in adipose-derived stem cells protects against UVB-induced photoaging in co-cultured human fibroblasts.

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